Effects of Bcl-2 modulation with G3139 antisense oligonucleotide on human breast cancer cells are independent of inherent Bcl-2 protein expression

We have investigated the effects of transient Bcl-2 down-regulation induced by the Bcl-2 antisense oligodeoxynucleotide (ODN) G3139 (Genta Incorporate d) in high Bcl-2 protein expressing, estrogen receptor (ER) positive MCF-7 and low Bcl-2 expressing, ER negative MDA435/LCC6 human breast cancer cells . Treatment with Bcl-2 antisense ODN in vitro caused > 80% reduction of Bcl -2 protein levels in a sequence specific manner for both cell lines. Maximu m mRNA reduction was achieved within 24 h of the first antisense ODN exposu re whereas full protein down-regulation required antisense exposure over 48 h. This Bcl-2 reduction was associated with 80-95% loss of viable cells co mpared to untreated cells. Similar cytotoxic effects were observed in both cell lines despite a nine-fold intrinsic difference in Bcl-2 protein expres sion suggesting that the relative degree of down-regulation of Bcl-2 is mor e important than the absolute reduction. Cell death associated with G3139 e xposure exhibited properties indicative of apoptosis such as mitochondrial membrane depolarization and caspase activation. Combined treatment with G31 39 and cytotoxic agents resulted in additive cytotoxicity in both cell line s. However, under most conditions studied, the direct cytotoxic activity of G3139 antisense was not synergistic with the cytotoxic agents. These resul ts suggest that while Bcl-2 clearly constitutes an attractive therapeutic t arget due to its role in regulating apoptosis in breast cancer cells, addit ional mechanisms are important in the control of apoptosis arising from exp osure to anticancer agents in vitro.